Colorimetric Detection

Colorimetric Detection of Mercury Ions Based on Plasmonic Nanoparticles Jianjun Du , Lin Jiang , Qi Shao , Xiaogang Liu , Robert S. Marks , Jan Ma , and Xiaodong Chen *

1467wileyonlinelibrary.com© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

The development of rapid, specifi c, cost-effective, and robust tools in monitoring Hg 2 + levels in both environmental and biological samples is of utmost importance due to the severe mercury toxicity to humans. A number of techniques exist, but the colorimetric assay, which is reviewed herein, is shown to be a possible tool in monitoring the level of mercury. These assays allow transforming target sensing events into color changes, which have applicable potential for in-the-fi eld application through naked-eye detection. Specifi cally, plasmonic nanoparticle-based colorimetric assay exhibits a much better propensity for identifying various targets in terms of sensitivity, solubility, and stability compared to commonly used organic chromophores. In this review, recent progress in the development of gold nanoparticle-based colorimetric assays for Hg 2 + is summarized, with a particular emphasis on examples of functionalized gold nanoparticle systems with oligonucleotides, oligopeptides, and functional molecules. Besides highlighting the current design principle for plasmonic nanoparticle-based colorimetric probes, the discussions on challenges and the prospect of next-generation probes for in-the-fi eld applications are also presented.

Introduction 1. ......................................... 1468

Correlation between Colorimetric 2. Detection and Plasmonic Nanoparticles ...................................... 1468

Oligonucleotide-AuNP Probes 3. .............. 1469

Oligopeptide-AuNP Probes4. ....................1473

Functional Molecule-AuNP Probes 5. .........1474

Multiple Signal Amplifi cation 6. ................1477

Conclusion and Outlook 7. ....................... 1478

From the Contents

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DOI: 10.1002/smll.201200811

Dr. J. Du, Dr. L. Jiang, Q. Shao, Prof R. S. Marks, Prof. J. Ma, Prof. X. ChenSchool of Materials Science and EngineeringNanyang Technological University50 Nanyang Avenue, 639798, Singapore E-mail: chenxd@ntu.edu.sg

Prof. X. LiuDepartment of ChemistryNational University of Singapore3 Science Drive 3, 117543, SingaporeInstitute of Materials Research and EngineeringA ∗ Star, 3 Research Link, 117602, Singapore

Prof. R. S. MarksDepartment of Biotechnology EngineeringNational Institute for Biotechnology in the Negevand the Ilse Katz Center for Meso and Nanoscale Science and TechnologyBen-Gurion University of the NegevBeer-Sheva, 84105, Israel

1. Introduction

Mercury is one of the most toxic and dangerous heavy metal

elements to human health and environmental safety. Mer-

cury contamination is toxic and widespread through various

natural and anthropogenic processes, such as volcanic emis-

sions, mining, solid waste incineration, and combustion of

fossil fuels. [ 1 ] Subsequent bioaccumulation through the food

chain can lead to severe damage to many important organs

such as the brain and kidney, as well as deleterious effects

on human health in the forms of dysphoria, tremors, vege-

tative nerve functional disturbance and so on. [ 2 ] The World

Health Organization (WHO) standard for the maximum

allowable level of inorganic mercury in drinking water is no

more than 6 ppb (30 nM). [ 3 ] Thus, concerns over the safety

of drinking water, damage associated with mercury toxicity,

and its toxicology, have provided the motivation to develop

rapid, specifi c, cost-effective, and robust tools for monitoring

ionic mercury (Hg 2 + ), which is one of most stable and wide-

spread inorganic forms of mercury in the environment and

organisms.

Although conventional analytical approaches for Hg 2 +

analysis, such as atomic absorption spectroscopy, [ 4 ] induc-

tively coupled plasma mass spectrometry, [ 5 ] and selective

cold vapor atomic fl uorescence spectrometry [ 6 ] could pro-

vide qualitative and quantitative information, they are

rather costly, complex, time-consuming, non-portable, and

require specialized laboratories. One should note that these

methods give accurate and sensitive quantifi cation of the

total amount of metal present in the sample. However,

before using these analytical methods, environmental sam-

ples require laborious treatment to solubilize the metal ions

from the solid matrix (i.e., soils or sediments which are sinks

for released metals). Still, a simplifi cation of these tech-

niques would be a welcome event. [ 7 ] Therefore, a variety of

remarkable probes based on organic molecules, [ 8 ] polymeric

materials, [ 9 ] biomaterials, [ 10 ] semiconductor nanocrystals, [ 11 ]

and agglutination materials [ 12 ] have been developed for

Hg 2 + detection over the past decades, which normally test

Hg 2 + using optical and electrochemical signals. [ 13 ] However,

most of examples are not well suited for on-fi eld analysis

or rapid screening. Alternatively, colorimetric detection, a

method of determining the existence and concentration of

an analyte with the aid of an obvious color change is par-

ticularly attractive for point-of-use applications due to its

simple readout by the naked eye. In recent years, a variety

of organic chromophoric probes have been utilized in mon-

itoring metal ions, [ 14 ] anions, [ 15 ] thiols, [ 16 ] amino acids, [ 17 ]

nucleotides, [ 18 ] and explosives. [ 19 ] However, their sensitivity,

solubility, and assay range usually limit the application

of organic chromophoric probes. [ 20 ] A new fi eld has been

developing rapidly to provide a possible solution to the

unsatisfi ed criteria for colorimetric detection. Indeed, plas-

monic nanoparticle-based colorimetric detection is drawing

increasing attention, as it could overcome the diffi culties

mentioned above.

Plasmonic nanoparticles, such as gold nanoparticles

(AuNPs) and silver nanoparticles (AgNPs), are a class of

nanostructures whose optical properties are determined by

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their unique surface plasmon resonances (SPRs). [ 21 ] Unlike

propagating plasmons supported on a bulk metal surface,

nanoparticle plasmons are the collective oscillations of their

conduction electrons confi ned to nanoscale volumes upon

light irradiation (incident electromagnetic waves). Such res-

onance is sensitive to nanoparticle size, shape, interparticle

distance, and local dielectric environment. [ 22 ] So far, various

nanoparticles with different sizes and shapes have been

developed for diverse applications. Since they have good sta-

bility and low toxicity, [ 23 ] several AuNP-based colorimetric

assays have been designed, prepared, and widely applied in

detecting DNA, [ 24 ] enzyme activity, [ 25 ] proteins, [ 26 ] small mol-

ecules, [ 27 ] anions/metal ions, [ 28 ] and so on.

The purpose of the present review is to summarize the

progress in the design and application of AuNP-based colori-

metric assays for Hg 2 + . We begin with a brief discussion on

the correlation between colorimetric detection and plasmonic

nanoparticles. Then, we describe the general design principles

that govern the nanoparticle-based colorimetric assays for

Hg 2 + detection. They are classifi ed according to the type of

functional decorators on nanoparticles such as oligonucle-

otides, oligopeptides, and functional molecules ( Figure 1 ).

Furthermore, the combination of multiple signal amplifi ca-

tion together with AuNPs for enhanced sensitivity is intro-

duced. Lastly, we present our views on the future outlook of

plasmonic nanoparticle-based colorimetric detection for on-

fi eld applications.

2. Correlation between Colorimetric Detection and Plasmonic Nanoparticles

There are essentially two key components of a colorimetric

assay that affect the performance in selectivity, sensitivity,

response time, and signal-to-noise ratio. One is the recogni-

tion moiety that provides a selective/specifi c response to

the analyte, which relates to a wide range of organic or bio-

logical ligands/reactions. The other is the transducer moiety

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Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

Jianjun Du is a Research Fellow at the

School of Materials Science and Engineer-

ing, Nanyang Technological University

(Singapore). He received his BS degree

in Chemistry from Dalian University of

Technology (China) in 2004, and obtained his

PhD from Dalian University of Technology

(China) in 2010. He is presently working as

a postdoctoral fellow in the group of Prof.

Xiaodong Chen. His research is focused on

fl uorogenic and colorimetric probes based on

organic and inorganic materials.

Xiaodong Chen is a Singapore National

Research Foundation (NRF) Fellow and

Nanyang Assistant Professor at the School of

Materials Science and Engineering, Nanyang

Technological University (Singapore). He

received his BS degree (Hons) in Chemistry

from Fuzhou University (China) in 1999, MS

degree (Hons) in Physical Chemistry from

the Chinese Academy of Sciences in 2002,

and PhD degree (Summa Cum Laude) in

Biochemistry from University of Muenster

(Germany) in 2006. After his postdoctoral

fellow working at Northwestern University

(USA), he started his independent research

career at Nanyang Technological University in 2009. His research interests include the

integrated nano–bio interface and bio-inspired nanomaterials for energy conversion.

that translates detecting behavior into an eye-sensitive color

change in the range of 390 to 750 nm.

Plasmonic nanostructures have the ability to support

surface plasmons ( Figure 2 a) [ 21a ] that are generated by the

coupling between incident electromagnetic waves with the

conduction electrons in metal nanostructures. [ 29 ] These kinds

of plasmons normally occur in AuNPs and AgNPs with a size

range of 10-200 nm and result in an amplifi cation of the elec-

tromagnetic fi eld near the particle surface. By tailoring the

size, shape, and environment of metal nanostructures, the

SPR adsorption of nanostructures can be tuned. In addi-

tion, when nanostructures approach one another (i.e., when

they aggregate), the SPR bands are dramatically altered as a

result of the strong coupling between the localized SPRs of

the nanostructures. [ 21b , 21c ]

Many plasmonic nanostructures with various sizes,

shapes (rod, cube, disc, etc.), and structures (shell, core-

shell, cage, tube, etc.) have been developed, [ 30 ] which

enable the tailoring of their SPR adsorption. For instance,

the shape and size of AgNPs can be controlled either

photochemically by adjusting the wavelength of irradia-

tion or chemically by varying the pH of the reaction solu-

tion. [ 31 ] As a result, a large range of SPR absorption bands

were obtained from visible region to near infrared region

(Figure 2 b,c). For example, AuNPs with the diameter of

10-50 nm usually appear ruby red in the solution. When

AuNPs aggregate, a dramatic red-to-blue color change can

be clearly observed, since the SPR adsorption displays a

red shift, broadening and decreasing in intensity compared

with that of isolated nanoparticles. Among various plas-

monic nanoparticles, AuNPs are most suitable for naked-

eye colorimetric detection because changes between red

and blue are dramatic and readily detected by human

eyes, which lead to their extensive application.

There are six main advantages to AuNP-based colorimetric

detection compared to organic chromophoric probes: [ 32 ] 1) ease

of synthesis with high dispersity in aqueous media; 2) greater

absorption extinction coeffi cient (ca. 10 8 cm − 1 M − 1 for AuNPs)

compared to that of common organic dyes (ca. 10 5 cm − 1 M − 1 );

3) strong photostability; 4) ease of physical and chemical

functionalization through suitable surface chemistry; 5) a

large surface-to-volume ratio with unusual target-binding

properties, and; 6) alterable optical properties (e.g., SPR)

which directly relate to the size, shape, and composition of

the nanoparticles, and their interparticle distance. There-

fore, AuNP-based colorimetric assays have been successfully

designed and widely applied.

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Figure 1 . Schematic illustration of AuNPs based colorimetric probe.

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3. Oligonucleotide-AuNP Probes

Oligonucleotide-AuNP probes have been widely used in the

fi eld of bio-nanotechnology for diagnosis and therapy [ 24a , 33 ]

due to their unique chemical and physical properties. [ 34 ] For

instance, both quantitative and qualitative detection of various

targets through precise and reversible control of the aggre-

gation of nanoparticles have been utilized as powerful diag-

nostic platforms to detect target analytes such as proteins, [ 35 ]

polynucleotides, [ 33a , 36 ] metal ions, [ 37 ] and small molecules. [ 38 ]

In the following sections, we will discuss how the oligonucle-

otide-AuNP probes are designed for detecting Hg 2 + .

3.1. dsDNA-AuNP Probes

In general, mixing two kinds of AuNP probes functionalized

with complementary ssDNA sequences would result in the

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Figure 2 . (a) Schematic of plasmon oscillation for a sphere, showing the displacement of the conduction electron charge cloud relative to the nuclei. Reproduced with permission. [ 21a ] Copyright 2003, The American Chemical Society. (b,c) Size- and shape-tunable surface plasmon resonance spectra of various AgNPs. Reproduced with permission. [ 31 ]

polymeric network-like aggregation of AuNPs accompanied

by a red-to-blue color change because of the red shift of the

SPR adsorption band. [ 24a , 33a , 39 ] Interestingly and importantly,

this process is reversible upon heating, which can de-hybridize

dsDNA along with the recovery of the original absorption

and color. The melting profi les of the complementary duplex

DNA strands labeled on AuNPs exhibit an extraordinarily

sharp transition compared to that of free DNA strands. [ 40 ]

Dependent on thymine-Hg 2 + -Thymine (T) coordination

chemistry, the colorimetric assay for Hg 2 + was fi rst introduced

and subsequently developed by the Mirkin group based on a

complementary DNA-AuNP system with designed T-T mis-

matches. [ 41 ] As shown in Figure 3 a, two types of nanoparti-

cles aggregated together after the specially designed ssDNA

hybridized, causing a red-to-purple color change. After

raising the temperature to the melting temperature, T m , the

dsDNA de-hybridized, which made AuNP aggregates disso-

ciate reversibly along with a color change back to red. Among

environmentally relevant metal ions (Mg 2 + , Pb 2 + , Cd 2 + , Co 2 + ,

Zn 2 + , Fe 2 + , Ni 2 + , Fe 3 + , Mn 2 + , Ca 2 + , Ba 2 + , Li + , K + , Cr 3 + , Cu 2 + , and

Hg 2 + ), only Hg 2 + could raise the T m obviously (ca. 5 ° C) as

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shown in Figure 3 b. The sharp melting transition enhanced

the sensitivity and lowered the limit of detection (LOD)

visibly to 100 nM (20 ppb) in contrast to organic colorimetric

system (LOD: usually in ppm range). Initially, dsDNA-AuNP

probes were developed for studying the imperfect DNA tar-

gets (such as single base mismatch or mutations), [ 24a , 36 ] as it

is known that the T m is decreased in the presence of base

mismatches or mutations in DNA strands. [ 42 ] A point to note

is that complementary DNA strands together with T-T mis-

matches can form a duplex DNA, creating a T-Hg 2 + -T com-

plex, thanks to Hg 2 + , which selectively bridges two thymine

molecules in a stable manner. [ 43 ] Such a unique structure

can obviously increase the T m in such a way that the melting

analysis could be used for qualitative determination. One can

design a smart, tailored ssDNA-AuNP system, which could

provide the user with notable selectivity toward Hg 2 + without

signifi cant interference from other metal ions. In addition,

this system takes advantage of the sensitivity provided by

AuNPs and thus, together, these properties can be considered

an improvement over traditional probes for Hg 2 + detection.

Nonetheless, heating is needed in this assay for thermal dena-

turation of dsDNA and the temperature change should be

monitored carefully, which impedes fast sample detection in

practice.

Based on the initial work of the Mirkin group, Liu et al.

developed a system that was not only selective and sensitive

but also practical and convenient for colorimetric detection of

Hg 2 + at room temperature. [ 44 ] Their studies showed that the

extent of reduced T m was proportional to the amount of T-T

mismatch and therefore taking advantage of this fact, the T m

could even be further reduced below the operating ambient

temperature (23 ° C) when the T-T mismatch amount reached

8 (probes A ∗ , B, and C 7 : T m = 14.4 ° C, Figure 3 d). Generally,

in order to detect target oligonucleotides for the diagnosis of

gene-pathogenic diseases, a three-ssDNA system is now pre-

ferred [ 36 , 45 ] wherein two different probe ssDNAs have been

carefully designed and functionalized to AuNPs respectively

as probe A and B , and together they complementarily recog-

nize different parts of the third target ssDNA ( C ) by forming

stable dsDNA. It is inferred that T m would become lower

and lower along with an increase of base mismatches, and

that these probes could even fi nally not hybridize when the

T m is lower than the operating temperature. However, in the

case where the mismatches are all T-T type, probe C would

recognize and hybridize both particle probe A and B via the

formation of stable dsDNA in the exclusive case that Hg 2 +

is present in the solution at a given operating temperature.

Therefore, colorimetric detection of target Hg 2 + at room tem-

perature is achieved through the systematic control and opti-

mization of the amount of T-T mismatches within the probe

C and A / B system (Figure 3 c,d), which renders the heating

and T m monitoring steps redundant. Thus a rapid detection of

Hg 2 + is shown that is selective to a single metal ion and that

provides a good sensitivity at room temperature.

Furthermore, Takarada et al. demonstrated that T-T mis-

matches in different positions could lead to differences in

the change of T m (Figure 3 e). [ 46 ] Different positions of T-T

mismatches in dsDNA were studied at the distal end, penul-

timate, and antepenultimate positions. The results showed

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Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

Figure 3 . (a) Schematic representation of colorimetric detection of Hg 2 + using ssDNA-AuNPs and (b) their normalized melting curves of aggregates in the presence of various metal ions. Reproduced with permission. [ 41 ] (c) Schematic representation of colorimetric detection of Hg 2 + using ssDNA-AuNPs and (d) their normalized melting curves of solutions containing probes A (or A ∗ ), B, and the linker probes C 1-7 with varied T-T mismatches. Reproduced with permission. [ 44 ] Copyright 2008, The American Chemical Society, and; (e) schematic illustration of spontaneous aggregation of dsDNA-carrying AuNPs with T-T mismatch in terminal.

that T-T mismatches in different positions could lead to

varying changes of T m and thus different LODs could be

obtained. One of the best visual LODs reached was, at 0.5

μ M Hg 2 + , using a system with T-T mismatches at the ante-

penultimate position. This system exhibited good selectivity

to Hg 2 + in 1 min without having to control the temperature

or mask reagents. In media with high ionic strength, fully

matched dsDNA tethered on the surface of AuNPs can sta-

bilize nanoparticles to some extent, because of the steric

hindrance from dsDNAs. However, dsDNA with base mis-

matches in the terminal end would form branch-type struc-

tures, which cause much more steric hindrance and larger

entropic repulsion than that of the rigid structures, resulting

in much higher colloidal stability for dispersions in identical

saline solutions.

It is known that free, non-tethered ssDNA (without a

thiol modifi cation at its end) can also attract AuNPs and

form complexes because of the coordination between Au

and the nitrogen atoms in DNA bases. [ 47 ] However, dsDNA

cannot follow such a behavior because DNA bases are

concealed after the hybridization event, while the nega-

tive phosphate backbone is exposed. Based on these facts,

Yang et al. presented examples of a new system based

on colorimetric and fl uorescent dual sensing where Hg 2 +

was coordinated within a dye-tagged ssDNA with T-T

mismatch sequences ( Figure 4 ). [ 48 ] Fluorescein (FAM)

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was introduced as the fl uorescence source, as its emis-

sion at 518 nm has a good overlap with the SPR band of

AuNPs (13 nm) at 520 nm; therefore, more than 95%

quenching could be observed through a fl uorescence reso-

nance energy transfer (FRET) process. In the absence

of Hg 2 + , the dye-tagged ssDNA (sequence 2 , Figure 4 b)

that was absorbed on the surface of AuNPs via an electro-

static effect, protected the particles from aggregation at high

ionic strengths and the FAM’s fl uorescence was quenched at

the same time. In the presence of Hg 2 + , sequence 2 , rich in

thymine bases and labeled with FAM at the 3′-end, could

form dsDNA with sequence 4 through vehicular Hg 2 + and

subsequently detach from the surface of the AuNPs. As a

result, fl uorescence was returned, accompanied by a red-to-

blue color change owing to the non-crosslinking aggregation

of AuNPs. This example is similar to the molecular beacon

system consisting of a fl uorescent dye and a corresponding

quencher in both ends of ssDNA, which is used for trapping

DNA targets for tailored FRET-based detection. In Yang’s

system, however, only the dye labeled ssDNA is needed,

resulting in less laborious and more cost-effective synthesis

compared to the known molecular beacon system, since

the AuNPs themselves are good fl uorescence quenchers. In

addition, neither a covalent tether of AuNPs with oligonu-

cleotides nor a subsequent purifi cation is required, which

makes the system more effi cient for practical applications.

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Figure 4 . (a) Schematic depiction of colorimetric and fl uorescent sensing of Hg 2 + based on the free AuNPs and (b) structures of fl uorescein (FAM) and the FAM-labeled ssDNAs. Reproduced with permission. [ 48 ] Copyright 2008, The American Chemical Society.

Figure 5 . (a) Schematic representation of Hg 2 + stimulated aggregation of AuNPs and (b) TEM images of non-aggregated AuNPs stabilized by ssDNA in the presence NaClO 4 and the aggregated AuNPs after addition of Hg 2 + . Reproduced with permission. [ 49 ] Copyright 2008, Wiley.

Three types of DNA-AuNP modes

have been summarized at this point:

1) a AuNP-dsDNA-AuNP mode, wherein

both ends of a dsDNA are tethered

on different nanoparticles through

Au-S bonds; 2) a dsDNA-AuNP mode,

wherein only one side of the dsDNA is

tethered on the nanoparticle, and; 3) a

dsDNA/Au mode, wherein the DNA is

totally free. Furthermore, in the exclu-

sive case of the development of AuNPs

based Hg 2 + probes, one can summarize

the systems as follows: 1) compared to

a typical ssDNA tethered AuNP system,

the free ssDNA/AuNP system provides

a more cost-effective, simple, and rapid

determination for Hg 2 + when based on

a non-crosslinking aggregating mode,

which requires neither functionalization,

nor separation or purifi cation processes;

2) with careful design, the geometry and

space structure of T-rich ssDNA could

be transformed from a free and uncoiled

state to a specifi c hybridization structure

(like a hairpin structure) using Hg 2 + as a

bridge (T-Hg 2 + -T) and induce the non-

crosslinking-based colorimetric detection

without the need to record a temperature

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change. Compared to well-studied

ssDNA tethered AuNPs systems, free

ssDNA/AuNPs systems have been well

developed in recent years and show

great future potential, as described in

detail in the following section.

3.2. T n -ssDNA-AuNP Probes

Simple, fast, and wide-range systems

for the detection of Hg 2 + using T n -

ssDNA have been developed. For

instance, Willner et al. prepared T-rich

nucleic acid (5′-TTCTTTCTTCCCTT-

GTTTGTT-3′) for Hg 2 + detection

( Figure 5 ). [ 49 ] After treating AuNPs (13

nm) with ssDNA, NaClO 4 (100 mM)

was added into the solution to maintain

a high level of salinity. Under this condi-

tion, red color with an absorption band

at 520 nm was observed before addi-

tion of Hg 2 + , indicating good dispersion

of the nanoparticles. Upon addition of

Hg 2 + the solution turned blue, induced

by aggregation of AuNPs, which was

detected by the naked eye and also

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Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

Figure 6 . (a) Schematic representation of colorimetric detection of Hg 2 + based on an oligopeptide-AuNP system; (b) colorimetric detection of Hg 2 + based on simply mixing AuNPs and oligopeptides with cysteines at both terminals.

confi rmed by a red shift and broadened peak in the UV–vis

spectrum. Macroscopic changes were attributed to the for-

mation of the nucleic acid duplex-folded complex with the

help of Hg 2 + , and following release of ssDNA from AuNPs.

As a result, the system became so destabilized that AuNPs

aggregated. This probe showed good selectivity to Hg 2 +

among normal metal ions except for Pb 2 + , which could actu-

ally be masked using 2,6-pyridinedicarboxylic acid (PDCA),

and analysis for Hg 2 + could be done at concentration of

100 nM, with an LOD of 10 nM (2 ppb). In addition, Yang's

group demonstrated an example based on thrombin-binding

aptamer (TBA, 5′-GGTTGGTGTGGTTGG-3′), which real-

ized an Hg 2 + colorimetric assay with an LOD of 200 nM. [ 50 ]

These ssDNA-AuNP sensing systems further simplify the fab-

rication of the detecting systems and provide us with a wide

detection range for Hg 2 + . It is also found that ssDNA chains

with different sequences show varying performances (like

LOD) for Hg 2 + detection, which indicate that the amount and

the position of thymine are both key factors in the ssDNA-

AuNP probe design.

Since the amount of thymine in ssDNA or the length of

the T n -ssDNA is directly related to the sensitivity toward

Hg 2 + , Chang et al. focused on the quantitative study by

testing T n -ssDNA at different lengths (T 7 , T 33 , and T 80 ). [ 51 ]

It was shown that although the longer ssDNA did better in

stabilizing AuNPs in the same concentration, T 33 performed

better than either T 7 or T 80 in the detection of Hg 2 + . This may

be due to the fact that T 7 was too short to form folded struc-

ture. In contract to this, T 80 was considered too long to dis-

sociate from AuNPs. Li et al. found that the aptamer T 10 even

performed better with an LOD of 0.6 nM under optimum

condition (0.1 M HAc-NaAc, pH 4.0, 20 ° C). [ 52 ] Therefore,

an appropriate amount of thymine should be selected when

considering an equilibrium point between stabilizing AuNPs

in the absence of Hg 2 + and dissociating from AuNPs in the

presence of Hg 2 + .

The common paradigm is that fl uorescence signals are

more sensitive than absorption ones in sensing systems. How-

ever, it has been shown here that similar LODs for Hg 2 +

were obtained from both such signals by Li et al. using their

ssDNA-AuNP systems. [ 53 ] It is thought that some dye-tagged

dsDNA remained attached to the surface of AuNPs during

dissociation in the presence of Hg 2 + . Once T-rich ssDNA

bound with enough target Hg 2 + (high concentration) and

formed a dsDNA structure through T-Hg 2 + -T complexes,

increasing overall positive charges of dsDNA, some dye-

labeled dsDNA bound to the AuNPs and quench themselves,

before electrostatic interactions dominated and aggregation

of the dsDNA/AuNPs system happened. These aforemen-

tioned results provide an additional understanding of the

interactions between DNA and AuNPs in the presence of low

or high Hg 2 + concentrations.

Using a similar system involving T n -ssDNA-AuNP for

Hg 2 + detection, Tseng et al. found that the presence of Mn 2 +

could accelerate and enhance the aggregation of AuNPs

in high salt media. [ 54 ] It was found that addition of Mn 2 +

(0.2 mM) would not affect the stability of system but could

increase the sensitivity by 100-fold to achieve 10 nM (LOD)

in Hg 2 + detection. The added Mn 2 + could stabilize the folded

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structure of the T n -ssDNA-AuNP complex, which could facil-

itate the release of poly-T DNA from the particle surface and

speed up their aggregation. This thanks to the fact that the

metal ions of Mn 2 + coordinate with DNA bases and bind to

the phosphate backbone of DNA owing to their specifi c ionic

valence and coordination geometry.

4. Oligopeptide-AuNP Probes

Oligopeptides are the source of highly specifi c ligands ena-

bling the capture of specifi c targets through a combination

of non-covalent interactions like hydrogen bonding, ionic

bonding, and π – π stacking found in 3D cavities fi lled with

functional groups from amino acids lying at the inner or

outer surfaces. Unlike DNA bases, there are about 20 kinds

of natural amino acids, all of which can act as building blocks

in the peptide/protein synthesis. Therefore, multiple permu-

tations and combinations with different functions become

possible, where even a tripeptide, for instance, has 20 3 kinds

of possible sequences. Oligopeptides show some unique intri-

cacies, which could be utilized in AuNP-based colorimetric

systems, such as 1) hydrophobicity/hydrophilicity can be

regulated by adjusting the confi guration of hydrophobic and

hydrophilic amino acids; 2) multiplexed readouts can supply

more choices (e.g., the indol group in tryptophane fl uoresces

at 350 nm); [ 55 ] 3) unlike DNA strands, multiple options in

introducing more functional groups are possible, like car-

boxyls, aminos, hydroxyls, thiols, and so on; 4) in addition,

oligopeptides also exhibit a redox capability, which could be

used in the synthesis of oligopeptide-AuNPs. In the following

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Figure 7 . Schematic representation of colorimetric detection for Hg 2 + based on functional molecules decorated AuNPs.

section, we will discuss how oligopeptide-

AuNP probes work in Hg 2 + detection.

For example, Mandal et al. prepared

an oligopeptide-AuNP system (NH 2 -Leu-

Aib-Tyr-COONa) in situ by oligopeptide

reduction and stabilization to work as

a colorimetric assay for Hg 2 + detection

( Figure 6 a). [ 56 ] In the absence of Hg 2 + , it

showed an initial sharp and characteristic

SPR band at 527 nm. Upon addition of

4 ppm Hg 2 + , a red-to-purple color change

was observed, while an additional 8 ppm

Hg 2 + brought a new SPR band at 670 nm

accompanied by a red-to-blue color

change. This special oligopeptide exhib-

ited a good selectivity toward Hg 2 + among

other metalic ions such as Pb 2 + , Cu 2 + , Cd 2 + ,

Zn 2 + , and Ca 2 + . More recently, Naik et

474

al. developed a colorimetric method for

detecting metal ions using oligopeptide-AuNPs. [ 57 ] The oli-

gopeptide (-Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys-Pro-Ala-

Tyr-Ser-Ser-Gly-Pro-Ala-Pro-Pro-Met-Pro-Pro-Phe-) was

selected (pI = 3.9) and its functional groups did interact with

metal ions, showing obvious colorimetric responses to several

metal ions in addition to Hg 2 + within a 1 min time period,

exhibiting an LOD as low as 26 nM for Hg 2 + . Chen et al. proposed a new colorimetric Hg 2 + assay

composed of AuNPs and oligopeptides (Lys-Cys-Gly-Trp-

Gly-Cys) without any pre-chemical modifi cation of the

AuNPs (Figure 6 b). [ 32 ] Thanks to a strong affi nity of Hg 2 +

for cysteine groups in oligopeptides (found in nature as poi-

soning effects through mercury's high affi nity to thiol groups

in enzymes), a strong ionic selectivity was obtained. In order

to span a wide detecting range of Hg 2 + (from a nanomolar

to a millimolar level) that would encompass the USA Envi-

ronmental Protection Agency (EPA) standards for both

industrial wastewater (250 nM; 50 ppb) and drinking water

(10 nM; 2 ppb), one would imagine having an array of dif-

ferent amounts of oligopeptides based on the estimated

concentration of Hg 2 + . Adding linear oligopeptides with Cys

at both ends into a solution containing AuNPs will provoke

their aggregation, as evidenced by a color change from red

to blue. Hg 2 + preferentially reacts with thiolates because

Hg 2 + and Cys have a signifi cantly higher affi nity than that

of Au-Cys. Thus, the solution of AuNPs remains stable and

red, which avoids false positives due to spontaneous particle

aggregation.

The cases just described illustrate not only the poten-

tial of these systems but also the fact that multiplicity of

oligopeptides from a multitude of sequential combinations

would open the opportunity to create a large number of

applications.

5. Functional Molecule-AuNP Probes

One may imagine a variety of functional molecules, in

addition to oligonucleotides and oligopeptides, that could

www.small-journal.com © 2013 Wiley-VCH Verlag Gm

also be introduced into AuNP-based systems, which would

either stabilize or aggregate the nanopaticles via cova-

lent binding, electrostatic attraction, or repulsive interac-

tions. The functional molecules have the potential to be

superior to oligonucleotides and oligopeptides, through

simplicity, convenience, and multiplicity. Their design phi-

losophy and methodology in creating specifi c functional

interactions of molecule-target and molecule-AuNP rely

on surface chemistry, colloid chemistry, and coordination

chemistry as found in the organic chromogenic and fl uo-

rescent probes fi eld. The framework of decorator-AuNPs

includes: 1) coordination-based systems, in which specifi c

ligands are functionalized on the surface of AuNPs through

chemical grafting (like S-Au bonds), the electrostatic effect,

or physical absorption, and; 2) chemodosimeter-based sys-

tems, wherein the state of AuNPs would be affected by the

reaction.

5.1. Coordination based Functional Molecule-AuNP Probes

Thiol is one of the main functional groups for tethering to

the surface of AuNPs due to the strong Au-S bond. Func-

tional groups with O, N, and S atoms also play important

roles in ligands, such as in carboxyls, aminos, hydroxyls,

as well as thiol groups. Therefore, derivatives of mercapto

aliphatic acid become preferred decorators for an AuNP

system. As early as 2001, Tupp et al. employed 11-mer-

captoundecanoic acid (MUA)-capped AuNPs (13 nm) as

colorimetric sensors for heavy metal ions ( Figure 7 ). [ 58 ]

Aqueous suspensions of MUA-capped AuNPs displayed

intense SPR absorptions centred at 526 nm, which ren-

dered the solution red. Since the recognizing moiety just

involved carboxyl groups, this system showed comprehen-

sive chelation to several heavy metal ions, such as Pb 2 + ,

Cd 2 + , and Hg 2 + . Aggregation of the particles induced by

those metal ions yielded both a shift in the SPR band and

a substantial increase in long-wavelength Rayleigh scat-

tering, which in turn led to a visual color change from red

bH & Co. KGaA, Weinheim small 2013, 9, No. 9–10, 1467–1481

Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

Figure 8 . Schematic representation of Hg 2 + detection based on NTA-capped AuNPs.

to blue. Subsequently, Chang et al. used 3-mercaptopropi-

onic acid (MPA) for Hg 2 + detection. [ 59 ] Results showed that

this system exhibited a higher effi ciency as well as better

selectivity to Hg 2 + than Ca 2 + , Sr 2 + , Mn 2 + , Cr 3 + , and espe-

cially Pb 2 + and Cd 2 + . Moreover, in the presence of PDCA,

the LOD was further improved down to 100 nM, meaning

that a short chain of mercapto aliphatic acid performed

better in Hg 2 + sensing. Functional molecule-AuNP systems

circumvent complicated syntheses in producing oligonucle-

otides or oligopeptides.

Taking advantage of the greater thiophilic tendency of

Hg 2 + compared to other metal ions such as Pb 2 + , Cd 2 + , and

Cu 2 + , Kim et al. reported an attractive colorimetric sensor

for Hg 2 + detection through an S-Hg 2 + -S interaction in a

dithioerythritol (DTET)-modifi ed AuNP system. [ 60 ] In phos-

phate buffer (10 mM, pH 6.6), the DTET-AuNPs exhibited

an obvious colorimetric response from red (525 nm) to blue

(670 nm) upon addition of Hg 2 + . Only two interfering ions

were found: Cu 2 + and Ba 2 + led to a slow precipitation of

AuNPs (over several hours), which could be masked by the

addition of ethylene diamine tetraacetic acid (EDTA). The

system showed a linear correlation between Ex 670 nm/525 nm

(the ratio of absoprtion coeffi cients at 670 nm and 525 nm)

and Hg 2 + concentration from the LOD of 100 nM to 600 nM

(R 2 = 0.9975).

Furthermore, it was found that the combination of lig-

ands supplied better performance toward specifi c metal ion

detection. For instance, in an adenosine monophosphate

(AMP)/MPA-AuNP system, AMP helped AuNPs disperse

well in a highly saline solution due to electrostatic repulsion,

while MPA was good at Hg 2 + binding. [ 61 ] Masking agents like

PDCA are usually used to increase the selectivity in some

assays, however, inspired by a good selectivity of thymine

toward Hg 2 + , a single thymine-involved derivative combined

with a mercapto group was developed and functionalized on

AuNPs for targeting Hg 2 + (Figure 7 ). [ 62 ] This probe showed

excellent selectivity to Hg 2 + due to the advantage of T-Hg 2 + -T

coordination chemistry, as well as a nanomolar level of LOD

in tapwater samples without any masking agent.

© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheimsmall 2013, 9, No. 9–10, 1467–1481

Functional molecules can either stabi-

lize the dispersion of nanoparticles for a

red-to-blue sensing mode or, conversely,

induce the aggregation of nanoparti-

cles, which could be prevented by targets

for a blue-to-red sensing mode. Indeed,

Chen and Gao et al. found that 3-nitro-

1 H -1, 2, 4-triazole (NTA) could stabilize

AuNPs against the aggregation induced

by 2-amino-2-hydroxymethyl-propane-1,3-

diol (Tris) ( Figure 8 ). [ 63 ] In the presence

of Hg 2 + , the NTA selectively bound Hg 2 + ,

subsequently detached NTA from the

surface of AuNPs, leading to Tris-induced

aggregation of AuNPs. No noticeable color

changes were observed in the presence of

other metal ions at concentrations up to

100 mM in the absence of masking agents.

In the optimized conditions, the LODs of

7 nM and 50 nM were achieved by spectrophotometer and

direct visualization, respectively. On the contrary, pyridine,

4,4-dipyridyl (DPY) and thymine were all good inducers of

AuNP aggregation. [ 64 ] Among the more common metal ions

of Ca 2 + , Cd 2 + , Cr 3 + , Co 2 + , Cu 2 + , Fe 3 + , Hg 2 + , Mg 2 + , Mn 2 + , Ni 2 + ,

Pb 2 + , Zn 2 + and so on, only Hg 2 + completely inhibited pyri-

dine/DPY/thymine-induced nanoparticle aggregation via for-

mation of pyridine/DPY/thymine-Hg 2 + complexes. The LOD

of DPY reached as low as 3.0 ppb because of its high affi nity

to Hg 2 + , and 40 nM was the minimum level detected in tap-

water and spring water samples. [ 64a ] Thymine did even better,

for which an LOD of 0.4 ppb was obtained and recoveries of

88.0-112.5% were achieved in the application of Hg 2 + assays

in real water samples. [ 64c ]

5.2. Functional Chemodosimeter-AuNP Probes

In addition to the coordination interaction based strategies,

the concepts and methodology of chemodosimeters [ 65 ] can

also be applied to AuNP-based colorimetric systems. For

instance, thiourea derivatives have been developed as optical

chemdosimeters for Hg 2 + based on the Hg 2 + induced desul-

furization reaction with urea derivatives as products. [ 66 ] Con-

sidering different behaviours of urea and thiourea toward

AuNPs, Han et al. demonstrated a new type of unmodifi ed

AuNP system for Hg 2 + detection with the assistance of a

thiourea chemodosimeter. [ 67 ] Thiourea derivatives could

strongly adsorb on the surface of AuNPs (The log K f of

Au(SCN) n is ca. 16.98. [ 68 ] ), which transformed nanoparticles

from hydrophilic to hydrophobic and then induced their

aggregation. After desulfurization, the product of urea loses

this capbility, so that the AuNP solution remained stable and

red ( Figure 9 a).

The thiol group (–SH) is similar to the SCN group in

that it can also bind AuNPs strongly. The log K f of Hg(SCN) n

(ca. 21.8), [ 68 ] however, exceeds that of Au(SCN) n , therefore,

it is expected that only Hg 2 + among metal ions has the

ability to remove thiolates chemisorbed onto the surface

of AuNPs (the log K f values for Co 2 + , Zn 2 + , Cd 2 + , Ni 2 + , Pb 2 + ,

1475www.small-journal.com

J. Du et al.

1476

reviews

Figure 9 . (a) Schematic representation of Hg 2 + detection based on chemodosimeter-AuNPs system. Reproduced with permission. [ 67 ] ; and (b) schematic representation of proposed mechanism for the Hg 2 + induced colorimetric response of QA-AuNP. Reproduced with permission. [ 71 ] Copyright 2010, The American Chemical Society.

Mn 2 + , Fe 2 + , Fe 3 + , Cr 3 + , and Cu 2 + are 1.72, 2.0, 2.8, 1.76, 1.48,

1.23, 1.31, 4.64, 3.08, and 10.4, respectively [ 68 ] ). To prove

this strategy, Taki et al. modifi ed AuNPs with HS-EG 3 , in

which a red color with an absorption peak at 520 nm was

observed. [ 69 ] Added Hg 2 + broke the Au-S bonds prior to the

aggregation process of AuNPs and this drastic red-to-blue

color change only occurred with Hg 2 + , but not Cu 2 + , Co 2 + ,

Mn 2 + , Fe 3 + , Ni 2 + , Hg 2 + , Pb 2 + , Zn 2 + , or Cd 2 + . The extremely

high affi nity of the thiol group for Hg 2 + once again explains

this selective response. Later, Huang et al. studied AuNPs

modifi ed by alkanethiols with different chain lengths

(2-mercaptoethanol, 4-mercraptobutanol, 6-mercapto-

hexanol, and 11-mercaptoundecanol), and they found that

www.small-journal.com © 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinh

the system of 4-mercraptobutanol-func-

tionalized AuNPs showed best selec-

tivity and sensitivity toward Hg 2 + after

a systematic comparison. [ 70 ] Moreover,

the introduction of quaternary ammo-

nium ((11-mercapto-undecyl)-trimethyl-

ammonium) as a functionalizing agent

could increase water solubility of the

system to further bring better sensi-

tivity (Figure 9 b). [ 71 ] Natural molecules

with thiol groups, furthermore, are also

applied in the examples of Hg 2 + detec-

tion, which avoid complicated synthesis.

For instance, the introduction of cysteine

into a poly(diallyldimethylammonium)

chloride-reduced AuNP solution aggre-

gated the AuNPs and turned the solution

color to purple, however, only Hg 2 + can

prevent this behavior due to the con-

sumption of cysteine in the form of Hg 2 + -

cysteine complexes, in which the LOD

achieved 25 nM. [ 72 ]

An additional phenomenon could be

used whereby surface-deposited Hg 2 + on

AuNPs could be reduced by a reducing

agent, with mercury forming a solid

amalgam-like structure on the surface of

the AuNPs, leading to a decrease in SPR

band intensity. As an illustrative example,

Tseng et al. used Tween 20-capped AuNPs

for the rapid and homogeneous detection

of Hg 2 + /Ag + . [ 73 ] In this system, Tween 20

stabilized the particles against the high

ionic strength, and the citrate reduced the

Hg 2 + /Ag + to form Hg/Ag-Au alloys which

dislodged Tween 20 from the surface

of AuNPs and then induced the aggre-

gation of nanoparticles in highly ionic

media ( Figure 10 a). With respect to Hg 2 +

detection, a masking agent such as NaCl

(0.1 M) was introduced into the system

to mask the effect from Ag + , leading to

the precipitation of Ag + in the form of

AgCl (the solubility is 1.8 × 10 − 10 ). Under

optimum conditions (for Hg 2 + : 0.24 nM

Tween 20-AuNPs, 80 mM Na 3 PO 4 , and 0.1 M NaCl), when

the concentration of Hg 2 + increased gradually from 0 to

1000 nM, an obvious colorimetric change occurred, from

red to purple. There were good linear correlations between

Ex 650 nm/520 nm and Hg 2 + concentration over 200-600 nM (R 2

= 0.9944) in drinking water and 300–1000 nM (R 2 = 0.9977)

in sea water with LODs of 100 nM and 200 nM, respectively.

Inspired by a similar strategy, Chen et al. presented a colori-

metric method with a blue-to-red color change in the deter-

mination of Hg 2 + and Ag + and a mechanism of stabilizing the

AuNPs through a redox-formed metal coating with the help

of ascorbic acid (AA). In the presence of Hg 2 + or Ag + , the

metal ions could be reduced by AA to form an Hg/Ag-Au

eim small 2013, 9, No. 9–10, 1467–1481

Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

Figure 10 . Illustration of the mechanism of (a) Tween 20- and (b) ascorbic acid-AuNP system for Hg 2 + detection. Reproduced with permission. [ 73 , 74 ] ; Copyright 2010 and 2011, The American Chemical Society.

alloy coating on the surface of AuNPs, which prevented the

aggregation induced by self-assembling ligand N -acetyl-L-

cysteine (Figure 10 b). [ 74 ] The examples above are based on

the stronger oxidative ability of Hg 2 + and Ag + compared to

other metal ions.

6. Multiple Signal Amplifi cation

AuNP-based colorimetric systems with high sensitivity need

to be further explored to satisfy WHO standards. Therefore,

methods where dual or even multiple signals are generated

by coupling AuNPs with more sensitive signals as well as

special structures or techniques, such as fl uorescence, [ 48 ]

silver spot-based scattered light, [ 75 ] hyper-Rayleigh scat-

tering (HRS), [ 76 ] unique core-shell nanostructures (Au@

HgS), [ 77 ] as well as immunoreaction-based lateral fl ow

strip biocomponent (LFSB) technique [ 78 ] and cloud point

© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheimsmall 2013, 9, No. 9–10, 1467–1481

extraction (CPE) technique, [ 79 ] for aug-

menting signal sensitivity.

The sensitivity of AuNP-based colori-

metric assays for Hg 2 + detection using

selective binding complexes of T-Hg 2 + -T

usually could not satisfy the aforemen-

tioned requirements. Therefore, utilizing

a secondary amplifi cation based on cata-

lytic properties induced by nanoparticles,

such as with the subsequent reduction

of Ag + in the presence of hydroquinone

provided a signifi cantly increased sen-

sitivity. [ 75 ] Scattered light from silver

spots was quickly measured with a Veri-

gene Reader, in which the LOD reached

2 ppb ( Figure 11 a). In addition, the

measurement of scattered light was more

convenient. Ray et al. demonstrated

a rapid, easy, and reliable method for

Hg 2 + detection based on the nonlinear

optical (NLO) properties of MPA/HCys/

PDCA-functionalized AuNPs. [ 76 ] Hyper-

Rayleigh scattering (HRS) technique

can be used for monitoring NLO proper-

ties on the basis of light scattering. The

HRS intensity of functionalized AuNPs

varied with the addition of different

amounts of Hg 2 + , and the LOD could

reach 5 ppb and 10 ppm by monitoring

HRS and color change, respectively. Fur-

thermore, the unique core-shell nanos-

tructure (Au@HgS) was formed by the

addition of Hg 2 + into a AuNP-SH 2 com-

plex solution, in which Hg 2 + selectively

reacted with H 2 S on the surface of the

AuNPs to form HgS quantum dots on

the surface of the AuNPs. It was proved

that this nanostructure could increase

the SPR absorption due to the inter-

action of neighboring AuNPs and synergistic effects of

the AuNPs with HgS, which supported this system's good

LODs of 5 μ M and 0.486 nM by the naked eye and UV–vis

spectra, respectively. [ 77 ] Besides the duplicate signal-ampli-

fying methods, some other amplifi cation techniques could

be useful, such as immunoreaction-based LFSB technique

and cloud point extraction of small-sized AuNPs. Based on

the thymine-rich hairpin DNA-modifi ed AuNP system and

immunoreaction technique between digoxin and anti-dig-

oxin antibodies on the LFSB, only the addition of Hg 2 + can

form duplex structures between hairpin DNA and digoxin-

labeled DNA on the LFSB and produce a characteristic

red band on the test zone for visual detection of Hg 2 + . [ 78 ]

The sensitivity of this system can reach 0.1 nM, without

instruments, with good selectivity. Another example of

AuNP-based Hg 2 + detection is developed on the basis of

CPE, [ 79 ] in which the extraction of PDCA-capped AuNPs

(4 nm) into the Triton X-114 rich phase was so hard in the

1477www.small-journal.com

J. Du et al.

147

reviews

Figure 11 . (a) Depiction of the chip-based scanometric detection of Hg 2 + using DNA-AuNPs. Reproduced with permission. [ 75 ] Copyright 2008, The American Chemical Society; and (b) schematic representation of Hg 2 + -triggered cloud point extraction of MPA/HCys-AuNPs for Hg 2 + detection.

absence of Hg 2 + that it was still colorless after the extrac-

tion (Figure 11 b). In the presence of Hg 2 + , however, it

was easy to get red Triton X-114 with rich AuNPs. There-

fore, the visual and colorimetric detection of Hg 2 + in the

Triton X-114-rich phase was possible by the naked eye and

UV–vis spectrophotometer, respectively. The introduction

of MPA and HCys on the PDCA-capped AuNPs could fur-

ther improve LODs to 5 ppb by the naked eye and 2 ppb

by spectrophotometer, respectively, in optimum conditions

(60 ° C, 30 min). A linear range of 2-100 ppb (R 2 = 0.9975)

was obtained by measuring the absorption at 520 nm in

Triton X-114 phase.

7. Conclusion and Outlook

The application of AuNPs for colorimetric assays typically

relies on strong coupling between the target analyte and

nanoparticle, causing aggregation which, in turn, leads to

a dramatic color change in the solution. This colorimetric

“readout” circumvents the relative complexity inherent

8 www.small-journal.com © 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinhe

in conventional optical imaging/detec-

tion methodologies and thus may prove

to be suitable for low-cost point-of-

care diagnostics. This review has pre-

sented various AuNP probes for the

colorimetric detection of Hg 2 + based

on oligonucleotides, oligopeptides,

and functional molecules in aqueous

media. The key characteristics of each

example, like selectivity, LOD, response

time, and their applications in real

samples are summarized in Table 1 ,

which show competitive potential of

AuNP-based assays. Colorimetric detec-

tion techniques are already commonly

used for qualitative and even semi-

quantitative analysis in labs (like pH test

papers and temperature test papers) and

even daily life (like urine test papers and

pregnancy test strips). Though AuNPs

are stable, biocompatible, and easy to

prepare, store, functionalize in water,

and test in in-the-fi eld detecting appli-

cations, there still remain challenges to

their application for the quantitative

detection of environmental samples at

the required LOD. Recent advances in

smart-phone applications such as bar-

code recognition using cameras, sparked

interesting ideas of using such devices as

in-fi eld detectors for instant diagnosis

of blood or saliva, wherein merely a

droplet is required on the phone's touch

screen. Inspired by this concept, it is

envisioned that a smart phone with soft-

ware designed for colorimetric analysis could supply much

more accurate and quantitative information supporting the

qualitative judgement by the naked eye. Another example is

the colorimetric analysis of our exhaled breath through an

array of crossreactive plasmonic nanoparticles. It could be

a fast, reliable, and portable diagnosis for lung cancer that

could potentially provide early warnings to mitigate serious

damage. It is exciting to imagine the frontiers of such par-

adigm-shifting applications, and we believe that plasmonic

nanoparticle-based fi eld assays and diagnoses could bring us

a much more “colorful” and bright future.

Acknowledgements

We thank the fi nancial support from SERC-TSRP (1021520015) and the National Research Foundation of Singapore (CREATE Pro-gramme of Nanomaterials for Energy and Water Management and NRF-RF2009-04).

im small 2013, 9, No. 9–10, 1467–1481

Plasmonic Nanoparticle Colorimetric Detection of Mercury Ions

1479www.small-journal.com© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Table 1. Key characteristics of AuNP-based probes for colorimetric detection of Hg 2 + .

Probe LOD [nM] Selectivity Temperature/time Real Sample Reference

Oligonucleotide-AuNP Probes:

DNA-AuNPs by Mirkin et al. 100 Hg 2 + 47 ° C - [ 41 ]

DNA-AuNPs by Liu et al. 1000 Hg 2 + RT (21.3 ° C) - [ 44 ]

DNA-AuNPs by Takarada et al. 500 Hg 2 + RT and < 1 min - [ 46 ]

DNA attached AuPs by Yang et al. - Hg 2 + a) RT and 30 min Tap/lake

water

[ 48 ]

T-rich DNA AuNPs by Willner et al. 10 Hg 2 + a) RT and 20 min - [ 49 ]

T-rich DNA AuNPs by Yang et al. 200 Hg 2 + a) RT and 30 min Lake water [ 50 ]

T 33 DNA- AuNPs by Chang et al. 250 Hg 2 + RT and 10 min - [ 51 ]

T 10 DNA-AuNPs by Li et al. 0.6 Hg 2 + RT and 10 min Tap/lake

water

[ 52 ]

T-rich DNA AuNPs by Li et al. 780 Hg 2 + and Pb 2 + RT and 30 min - [ 53 ]

T 33 DNA-AuNPs by Tseng et al. 10 Hg 2 + a) RT and 5 min Pond water [ 54 ]

Oligopeptide-AuNP Probes:

Oligopeptide-AuNPs by Mandal et al. 2 × 10 4 Hg 2 + RT and 10 min - [ 56 ]

Oligopeptide-AuNPs by Naik et al. 26 Hg 2 + , Co 2 + , Pd 2 + , Pd 4 + , Pt 2 + RT and < 1 min - [ 57 ]

Oligopeptide-AuNPs by Chen et al. 10 Hg 2 + RT - [ 32 ]

Functional Molecule-AuNP Probes:

11-mercaptoundecanoic AuNPs

by Tupp et al.

- Pb 2 + , Cd 2 + , Hg 2 + RT - [ 58 ]

MPA-AuNPs by Chang et al. 100 Hg 2 + a) RT and < 60 min - [ 59 ]

DTET-AuNPs by Kim et al. 100 Hg 2 + a) RT and < 10 min - [ 60 ]

MPA/AMP-AuNPs by Yu and Tseng et al. 500 Hg 2 + RT and 30 min - [ 61 ]

Thymine-SH AuNPs by Chen et al. 2.8 Hg 2 + RT and 5 min Tap water [ 62 ]

Tris/NTA-AuNPs by Chen and Gao et al. 7 Hg 2 + RT and 1 h Lake water [ 63 ]

DPY-AuNPs by Zhong et al. 15 Hg 2 + RT and30 min Tap/Spring

water

[ 64a ]

Pyridine-AuNPs by Tian et al. 55 Hg 2 + RT and < 10 min Tap water [ 64b ]

Thymine-AuNPs by Chen et al. 2 Hg 2 + RT and30 min Tap water [ 64c ]

Thiourea-AuNPs by Han et al. 193 Hg 2 + and Ag + RT and 3 min - [ 67 ]

HS-EG 3 -AuNPs by Taki et al. - Hg 2 + RT and < 30 min - [ 69 ]

4-Mercraptobutanol AuNPs

by Huang et al.

500 Hg 2 + RT and 30 min River water [ 70 ]

MTA-AuNPs by Wang and Jiang et al. 30 Hg 2 + RT - [ 71 ]

Cysteine-AuNPs by Zhao, He, Zhang et al. 25 Hg 2 + RT Drinking

water

[ 72 ]

Tween 20-AuNPs by Tseng et al. 100 Hg 2 + and Ag + RT and 5 min Drinking/sea

water

[ 73 ]

AA-AuNPs by Chen et al. 5 Hg 2 + and Ag + RT and 3 min Drinking/tap

water

[ 74 ]

Multiple SignalAmplifi cation Probes:

Silver spot based scattered

light-AuNPs by Mirkin et al.

10 Hg 2 + 34 ° C and 2 h Lake water [ 75 ]

HRS-AuNPs by Ray et al. 25 Hg 2 + a) RT and 6-7 min - [ 76 ]

Au@HgS nanostructure by

Wang, Wu et al.

0.486 Hg 2 + RT and 1.5 h Pond water [ 77 ]

Immunoreation-AuNPs by He, Liu et al. 0.1 Hg 2 + RT and 20 min Tap and River

water

[ 78 ]

Cloud point extraction-AuNPs by Liu et al. 10 Hg 2 + a) 60 ° C and 30 min - [ 79 ]

a) Using PDCA as masking agent.

small 2013, 9, No. 9–10, 1467–1481

J. Du et al.

14

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Received: April 16, 2012Published online: September 7, 2012

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